Urazaev A K, Grossfeld R M, Fletcher P L, Speno H, Gafurov B S, Buttram J G, Lieberman E M
Department of Physiology, The Brody School of Medicine of East Carolina University, Greenville, NC 27858, USA.
Neuroscience. 2001;106(1):237-47. doi: 10.1016/s0306-4522(01)00270-6.
Early physiological and pharmacological studies of crayfish and squid giant nerve fibers suggested that glutamate released from the axon during action potential generation initiates metabolic and electrical responses of periaxonal glia. However, more recent investigations in our laboratories suggest that N-acetylaspartylglutamate (NAAG) may be the released agent active at the glial cell membrane. The investigation described in this paper focused on NAAG metabolism and release, and its contribution to the appearance of glutamate extracellularly. Axoplasm and periaxonal glial cell cytoplasm collected from medial giant nerve fibers (MGNFs) incubated with radiolabeled L-glutamate contained radiolabeled glutamate, glutamine, NAAG, aspartate, and GABA. Total radiolabel release was not altered by electrical stimulation of nerve cord loaded with [(14)C]glutamate by bath application or loaded with [(14)C]glutamate, [(3)H]-D-aspartate or [(3)H]NAAG by axonal injection. However, when radiolabeled glutamate was used for bath loading, radiolabel distribution among glutamate and its metabolic products in the superfusate was changed by stimulation. NAAG was the largest fraction, accounting for approximately 50% of the total recovered radiolabel in control conditions. The stimulated increase in radioactive NAAG in the superfusate coincided with its virtual clearance from the medial giant axon (MGA). A small, stimulation-induced increase in radiolabeled glutamate in the superfusate was detected only when a glutamate uptake inhibitor was present. The increase in [(3)H]glutamate in the superfusion solution of nerve incubated with [(3)H]NAAG was reduced when beta-NAAG, a competitive glutamate carboxypeptidase II (GCP II) inhibitor, was present.Overall, these results suggest that glutamate is metabolized to NAAG in the giant axon and its periaxonal glia and that, upon stimulation, NAAG is released from the axon and converted in part to glutamate by GCP II. A quisqualate- and beta-NAAG-sensitive GCP II activity was detected in nerve cord homogenates. These results, together with those in the accompanying paper demonstrating that NAAG can activate a glial electrophysiological response comparable to that initiated by glutamate, implicate NAAG as a probable mediator of interactions between the MGA and its periaxonal glia.
对小龙虾和鱿鱼巨神经纤维的早期生理学和药理学研究表明，动作电位产生期间轴突释放的谷氨酸会引发轴周神经胶质细胞的代谢和电反应。然而，我们实验室最近的研究表明，N-乙酰天冬氨酰谷氨酸（NAAG）可能是在神经胶质细胞膜上起作用的释放因子。本文所述的研究聚焦于NAAG的代谢和释放，以及它对细胞外谷氨酸出现的作用。从内侧巨神经纤维（MGNF）收集的轴浆和轴周神经胶质细胞胞质，在与放射性标记的L-谷氨酸一起孵育后，含有放射性标记的谷氨酸、谷氨酰胺、NAAG、天冬氨酸和γ-氨基丁酸。通过浴槽给药将[(14)C]谷氨酸加载到神经索中，或通过轴突注射加载[(14)C]谷氨酸、[(3)H]-D-天冬氨酸或[(3)H]NAAG后，电刺激神经索并不会改变总的放射性标记释放。然而，当使用放射性标记的谷氨酸进行浴槽加载时，刺激会改变超滤液中谷氨酸及其代谢产物之间的放射性标记分布。NAAG是最大的部分，在对照条件下约占回收的总放射性标记的50%。超滤液中放射性NAAG的刺激增加与其从内侧巨轴突（MGA）的实际清除同时发生。只有当存在谷氨酸摄取抑制剂时，才检测到超滤液中放射性标记的谷氨酸有小幅的、刺激诱导的增加。当存在竞争性谷氨酸羧肽酶II（GCP II）抑制剂β-NAAG时，与[(3)H]NAAG一起孵育的神经的超融合溶液中[(3)H]谷氨酸的增加会减少。总体而言，这些结果表明，谷氨酸在巨轴突及其轴周神经胶质细胞中代谢为NAAG，并且在受到刺激时，NAAG从轴突释放，并通过GCP II部分转化为谷氨酸。在神经索匀浆中检测到对quisqualate和β-NAAG敏感的GCP II活性。这些结果，连同随附论文中表明NAAG可以激活与谷氨酸引发的反应相当的神经胶质电生理反应的结果，表明NAAG可能是MGA与其轴周神经胶质细胞之间相互作用的介质。
